PROJECT SUMMARY More than 65% of women entering pregnancy in the US are overweight or obese. Obesity in pregnancy predisposes the offspring to obesity, and cardiovascular and metabolic disorders, thus initiating a ?vicious cycle? of obesity and its health-related consequences in subsequent generations even in the absence of further intrauterine stressors. The worldwide epidemic of obesity and cardiovascular and metabolic diseases, therefore, is not only a result of the sedentary lifestyle or poor diet; it is also a consequence of a ?developmental program? switched on as a result of an adverse in utero environment. The absence of therapeutic strategies to prevent developmental programming is a consequence of our lack of knowledge of the mechanisms whereby maternal obesity affects the health of future generations. The placenta is a crucial determinant of healthy fetal growth and development, but the consequences of obesity for placental health are only now beginning revealed. Data from our laboratory have shown an accumulation of inflammatory cytokine TNF? in the placentas of female fetuses is associated with an activation of pro-inflammatory transcription factor nuclear factor ?appa B (NF?B). Inflammation is tightly regulated by autophagy, a cellular recycling process, and unresolved inflammation has been associated with a disruption of autophagy, which, in turn, leads to cardiovascular and metabolic diseases. Our recent data suggest that placentas of both male and female offspring from obese women show inhibition of autophagy. However, there are significant sex differences in the mechanisms. In the placentas of males, inhibition of autophagy is associated with a decrease in critical autophagy coordinating transcription factor EB. In the placentas of females, in contrast, inhibition of autophagy is linked to inflammation via Rubicon (RUN domain and cysteine-rich domain containing Beclin1- interacting protein), a newly identified negative regulator of autophagy. We found an increase in Rubicon expression and Beclin 1 binding in the placentas of females of obese women with this phenomenon not seen in placentas of males. Our data suggest that these processes are, at least in part, regulated by TNF?-induced inflammation. Here we hypothesize that in response to inflammation generated in utero by maternal obesity, an increase in placental Rubicon expression occurs causing changes in its protein-protein interaction and thereby inhibiting placental autophagy. We will address this hypothesis with three specific aims: 1). Identify the mechanisms that regulate expression of Rubicon and its binding to Beclin1 in the human placenta with maternal obesity. 2). Determine the molecular basis relating Rubicon to the inhibition of autophagy in the placenta with obesity. 3). Identify sex-dependent mechanisms that regulate Rubicon signaling in the placenta with maternal obesity. This study will provide mechanistic insights that will systematically and rigorously fill these gaps and thereby open the opportunity for the development of Rubicon-based therapies to address the adverse consequences of maternal obesity for offspring, a major unmet public health need.